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Genetic Perturbation Platform

The Genetic Perturbation Platform, formerly known as the RNA interference (RNAi) Platform, supports functional investigations of the mammalian genome that can reveal how genetic alterations lead to changes in phenotype.

To enable those investigations, the platform develops technologies for perturbing genes, including libraries of CRISPR/Cas9 constructs, short hairpin RNAs (shRNAs), and open reading frames (ORFs) to edit, knockdown, or overexpress genes, respectively.

In addition to developing technologies for perturbing genes, the platform also works to improve the effectiveness of the techniques, enhance delivery methods, and create infrastructure and resources to enable their use on large and small scales. Importantly, the team also assists collaborators in experimental planning and execution, helping them to choose the best model system and most appropriate readout to assess effects of the chosen perturbation.

The platform’s scope has grown significantly since its inception, when it grew out of The RNAi Consortium (TRC), a collaborative effort of six academic research institutions and five leading life sciences organizations. The Genetic Perturbation Platform and TRC worked as an integrated team to develop the materials and technology to enable and enhance RNAi as a tool for mammalian genetic screening. The materials and knowledge generated by this team were made available to the entire scientific community. The platform has since expanded to include varied forms of perturbing gene function, including a significant focus on CRISPR/Cas9 constructs.

The Genetic Perturbation Platform is directed by David Root, a physical chemist with significant experience in cell-based screening and building lentiviral-based libraries.

The platform's major activities include:

Library production

Platform scientists are creating genome-scale libraries of RNAi reagents, targeting virtually all human and mouse genes, which are carried in lentiviral vectors that allow this library to be introduced into a wide range of cell types. They have also developed a streamlined production process for rapid expansion of the library.

The platform is also building genome-scale libraries of CRISPR/Cas9 reagents, to knock out genes or perform genome editing.

In 2011 the RNAi Platform, in collaboration with the Broad Institute Cancer Program and the Center for Cancer Systems Biology at the Dana-Farber Cancer Institute, made publicly available the human ORFeome library V8.1, providing an additional tool for manipulating the human genome.

Library screening

Platform scientists are developing and optimizing methods for using the library in high-throughput screening. This has enabled the first arrayed RNAi genetic screens by lentiviral infection as well as pooled screening approaches.

Library performance

Platform scientists are evaluating the performance of the entire library using quantitative PCR to measure knockdown of the target transcript, to create the first fully-validated lentiviral RNAi libary. They are also testing and optimizing the performance of the library under different conditions, including different cell lines, timepoints, and multiplicity of infection (MOI).

Library development

New versions of the library are being developed, as well as new methods to screen large subsets of the existing library.

Informatics

Informatics scientists in the platform are developing archival and analytical tools necessary to ensure the utility of this library for all scientists.